Individual variation in the milk yield response to bovine somatotropin in dairy cows

M. T. Rose, T. E. C. Weekes and P. Rowlinson (2004). Individual variation in the milk yield response to bovine somatotropin in dairy cows. Journal of Dairy Science, 87(7), 2024-2031. Sponsorship: industry RAE2008

Production of polyclonal antibodies directed to recombinant methionyl bovine somatotropin

The administration of recombinant methionyl bovine somatotropin (rMbST) to dairy cows to increase milk yield remains a common practice in many countries including the USA, Brazil, Mexico, South Africa and Korea, whereas it has been forbidden within the European Union (EU) since 1999. A rapid screening immunoanalytical method capable of the unequivocal determination of rMbST in milk would be highly desirable in order to effectively monitor compliance with the EU-wide ban for home-made or imported dairy products. For decades, the production of specific antibodies for this recombinant isoform of bovine somatotropin (bST) has remained elusive, due to the high degree of sequence homology between both counterparts (e.g. methionine for rMbST in substitution of alanine in bST at the N-terminus). In this study, we compared several immunizing strategies for the production of specific polyclonal antibodies (pAbs), based on the use of the full-length recombinant protein, an rMbST N-terminus peptide fragment and a multiple antigen peptide (MAP) which consists of an oligomeric branching lysine core attached to the first two N-terminus amino acids of rMbST, methionine and phenylalanine (MF-MAP). The immunization with KLH-conjugated MF-MAP led to the production of the pAb with the highest rMbST/bST recognition ratio amongst the generated battery of antibodies. The pAb exhibited a specific binding ability to rMbST in a competitive antigen-coated ELISA format, which avidity was further improved after purification by rMbST N-terminus peptide-based affinity chromatography. These results suggest that immunodiscrimination between structurally related proteins can be achieved using immuno-enhanced immunogens such as MAPs. © 2012 Elsevier B.V.

Muscarinic M1 and M3 Receptors,IP3 and cGMP Functional Regulation in Streptozotocin Induced Diabetic Rats: Insulin and Somatotropin Induced Rejuvenation as a Function of Age

The present study describes that acetylcholine through muscarinic Ml and M3 receptors play an important role in the brain function during diabetes as a function of age. Cholinergic activity as indicated by acetylcholine esterase, a marker for cholinergic function, decreased in the brain regions - the cerebral cortex, brainstem and corpus striatum of old rats compared to young rats. in diabetic condition, it was increased in both young and old rats in cerebral cortex, and corpus striatum while in brainstem it was decreased. The functional changes in the muscarinic receptors were studied in the brain regions and it showed that muscarinic M I receptors of old rats were down regulated in cerebral cortex while in corpus striatum and brainstem it was up regulated. Muscarinic M3 receptors of old rats showed no significant change in cerebral cortex while in corpus striatum and brainstem muscarinic receptors were down regulated. During diabetes, muscarinic M I receptors were down regulated in cerebral cortex and brainstem of young rats while in corpus striatum they were up regulated. In old rats, M I receptors were up regulated in cerebral cortex, corpus striatum and in brainstem they were down regulated. Muscarinic M3 receptors were up regulated in cerebral cortex and brainstem of young rats while in corpus striatum they were down regulated. In old rats, muscarinic M l receptors were up regulated in cerebral cortex, corpus striatum and brainstem. In insulin treated diabetic rats the activity of the receptors were reversed to near control. Pancreatic muscarinic M3 receptor activity increased in the pancreas of both young and old rats during diabetes. In vitro studies using carbachol and antagonists for muscarinic Ml and M3 receptor subtypes confirmed the specific receptor mediated neurotransmitter changes during diabetes. Calcium imaging studies revealed muscarinic M I mediated Ca2 + release from the pancreatic islet cells of young and old rats. Electrophysiological studies using EEG recording in young and old rats showed a brain activity difference during diabetes. Long term low dose STH and INS treated rat brain tissues were used for gene expression of muscarinic Ml, M3, glutamate NMDARl, mGlu-5,alpha2A, beta2, GABAAa1 and GABAB, DAD2 and 5-HT 2C receptors to observe the neurotransmitter receptor functional interrelationship for integrating memory, cognition and rejuvenating brain functions in young and old. Studies on neurotransmitter receptor interaction pathways and gene expression regulation by second messengers like IP3 and cGMP in turn will lead to the development of therapeutic agents to manage diabetes and brain activity.From this study it is suggested that functional improvement of muscarinic Ml, M3, glutamate NMDAR1, mGlu-5, alpha2A, beta2, GABAAa1 and GABAB, DAD2 and 5-HT 2C receptors mediated through IP3 and cGMP will lead to therapeutic applications in the management of diabetes. Also, our results from long term low dose STH and INS treatment showed rejuvenation of the brain function which has clinical significance in maintaining healthy period of life as a function of age.

Effect of recombinant bovine somatotropin (bST) on follicular population and on in vitro buffalo embryo production

The objective of this study was to evaluate the effect of bovine somatotropin (bST) on ovarian follicular population in buffalo heifers and its influence on oocyte quality, recovery rates and in vitro embryo production. We tested the hypothesis that bST treatment in buffalo females submitted to an ovum pick-up (OPU) program Would improve the number of follicles recruited, oocyte quality and in vitro embryo production. A total of 10 heifers were assigned into two treatment groups: group bST (n = 5; receiving 500 mg of bST in regular intervals) and control group (n = 5; without additional treatment). Both groups were subjected to OPU sessions twice a week (every 3 or 4 days), for a total of 10 sessions per female, although due to procedural problems, only the first five OPU sessions produced embryos. The number of follicles and the diameters were recorded at all OPU sessions. The harvested oocytes were counted and classified according to their quality as either A, B, C, D or E, with A and B considered good quality. Cleavage and blastocyst production rates were evaluated 2 and 7 days after in vitro fertilization, respectively. The bST treatment increased the total number of antral follicles (> 3 mm in diameter; 12.2 compared with 8.7: p, < 0.05) and of small antral follicles (< 5 mm; 9.1 compared with 6.5; p < 0.05) per OPU session. The bST also tended to increase the number of oocytes recovered per session (5.2 compared with 4.1; p = 0.07), and enhanced the percentage of good quality oocytes (48.8% compared with 40.6%; p = 0.07), bST showed no effect on cleavage and blastocyst production rates (p > 0.05). The significant effects of performing repeated OPU sessions were decreasing the follicular population (p < 0.001) as well as the number of follicles aspirated (p < 0.001), and oocytes recovered (p < 0.02). In conclusion, bST treatment improves the follicular population, demonstrating its possible application in buffalo donors submitted to OPU programs. (c) 2008 Elsevier B.V. All rights reserved.

Plasma progesterone concentration in beef heifers receiving exogenous glucose, insulin, or bovine somatotropin

Three experiments were conducted to evaluate plasma concentrations of glucose, insulin, IGF-I, and progesterone (P4) in pubertal beef heifers receiving exogenous glucose, insulin, or sometribove zinc. All heifers used had no luteal P4 synthesis but received a controlled internal drug-releasing device containing 1.38 g of P4 to estimate treatment effects on hepatic P4 degradation. In Exp. 1, 8 pubertal, nulliparous Angus x Hereford heifers (initial BW = 442 +/- 14 kg; initial age = 656 +/- 7 d) were randomly assigned to receive, in a crossover design containing 2 periods of 10 h, intravenous (i.v.) infusions (10 mL) of insulin (1 mu g/kg of BW; INS) or saline (0.9%; SAL). Treatments were administered via jugular venipuncture in 7 applications (0.15 mu g insulin/kg BW per application) 45 min apart (from 0 to 270 min). Blood samples were collected immediately before each infusion as well as at -120, -60, 330, 390, and 450 min relative to the first infusion. Heifers receiving INS had greater (P < 0.01) plasma insulin, reduced (P <= 0.04) plasma glucose and IGF-I, and similar (P = 0.62) plasma P4 concentrations compared with SAL heifers. In Exp. 2, the same heifers were assigned to receive, in a similar experimental design as Exp. 1, i.v. infusions (10 mL) of 1) insulin (1 mu g/kg BW) and glucose (0.5 g/kg BW; INS+G) or 2) SAL. Heifers receiving INS+G had greater (P <= 0.02) plasma insulin, glucose, and P4 but reduced (P = 0.01) plasma IGF-I concentrations compared with SAL heifers. In Exp. 3, the same heifers were assigned to receive, in a crossover design containing 2 periods of 14 d, subcutaneous (s.c.) injections of 1) 250 mg of sometribove zinc (BST) or 2) SAL. Blood samples were collected 3 h apart (0900, 1200, 1500, and 1800 h) from heifers on d 6, 8, and 10 relative to treatment administration (d 1). Heifers receiving BST had greater (P < 0.01) plasma glucose and IGF-I and similar (P >= 0.67) plasma insulin and P4 concentrations compared with SAL heifers. Results from this series of experiments suggested that concurrent increases in glucose and insulin are required to reduce hepatic catabolism and increase plasma concentrations of P4 in bovine females.

INFLUENCE OF DIETARY-PROTEIN INTAKE AND RECOMBINANT HUMAN SOMATOTROPIN ADMINISTRATION ON GROWTH AND BODY-COMPOSITION OF JUVENILE TAMBACU (A PIARACTUS-MESOPOTAMICUS X COLOSSOMA-MACROPOMUM CROSS)

This experiment was undertaken to study the interaction between level of dietary protein and recombinant human somatotropin (rhGH) administration on performance and body composition of juvenile tambacu (a crossbred Brazilian fish). A total of 72 juvenile tambacu, initially weighing and measuring (mean +/- s.e.m.) 23 +/- 2 g and 9 +/- 0.5 cm, respectively, were randomly divided into 18 groups of 4 fish each. Water temperature was 28 degrees C. Triplicate groups received one of two levels of dietary protein (15 and 30% as fed basis) and one of 3 doses of rhGH (0, 2 and 4 mu g/g) via intraperitoneal injection twice a week for 6 weeks, using a randomized complete block design. Somatotropin was noted to stimulate linear and body weight gain. The higher protein level supported increased growth in weight and length, but there was no interaction between protein level and rhGH dose for either parameter. Protein efficiency ratio and percentage protein deposited showed higher values on diets containing 15% protein. Somatotropin treatment did not significantly affect body composition, but there was a trend towards improved protein retention and reduced carcass lipid. In conclusion, the results of this experiment suggest that rhGH is able to stimulate linear gain in tambacu.

Effects of dominant follicle aspiration and treatment with recombinant bovine somatotropin (BST) on ovarian follicular development in Nelore (Bos indicus) heifers

Follicle ablation has been recognized as an efficient method of follicular wave synchronization. Treatment with recombinant bovine somatotropin (BST) has been shown to enhance follicular development in <(Bos taurus)under bar>. This experiment assessed the effects of these treatments in Nelore (<(B. indicus)under bar>) heifers. Eight cycling Nelore heifers were randomly assigned to 3 different treatments. on Day 2 of a synchronized cycle (Day 0 = day of ovulation), heifers assigned to Treatments 1 and 2 received 2 mL of saline, whereas heifers assigned to Treatment 3 received 320 mg of BST. on Day 5, the first-wave dominant follicle was ablated by ultrasoundguided transvaginal aspiration in heifers in Treatments 2 and 3, and all heifers received an injection of prostaglandin on Day 11. Aspiration of the dominant follicle advanced and synchronized (P < 0.05) the day of second-wave emergence (6.9 +/- 0.1 vs. 8.4 +/- 0.4) and the day of the pre-wave FSH peak (6.0 +/- 0.0 vs. 6.9 +/- 0.4), and increased FSH peak concentrations (381 +/- 21 vs. 292 +/- 30; pg/mL; P < 0.01). Recombinant bovine somatotropin treatment caused a two-fold increase in plasma insulin-like growth factor-I (IGF-I) concentrations (P < 0.001) and resulted in a 36% increase in the number of small follicles (<5 mm; P < 0.001) compared with saline-treated heifers. In summary, in agreement with previous reports on <(B. taurus)under bar>, dominant follicle aspiration synchronized ovarian follicular development, and BST treatment increased peripheral concentrations of IGF-I in Nelore heifers. Recombinant bovine somatotropin also increased the number of small follicles, but this response appeared to be inferior to that reported for <(B. taurus)under bar>. (C) 2000 by Elsevier B.V.

Effects of flunixin meglumine, recombinant bovine somatotropin and/or human chorionic gonadotropin on pregnancy rates in Nelore cows

The objective was to compare pharmacological strategies aiming to inhibit prostaglandin F2 alpha (PGF(2 alpha)) synthesis (flunixin meglumine; FM), stimulate growth of the conceptus (recombinant bovine somatotropin; bST) and progesterone (P(4)) synthesis (human chorionic gonadotropin; hCG), as well as their combinations, regarding their ability to improve pregnancy rates in beef cattle. Lactating Nelore cows (N = 975), 35 to 70 days postpartum, were synchronized and inseminated by timed artificial insemination (TAT) on Day 0. on Day 7, cattle were allocated into eight groups and received one of the following treatments: saline (S) on Days 7 and 16 (Group Control); S on Day 7 and FM on Day 16 (Group FM); bST on Day 7 and S on Day 16 (Group bST); bST on Day 7 and FM on Day 16 (Group bST + FM); hCG on Day 7 and S on Day 16 (Group hCG); hCG on Day 7 and FM on Day 16 (Group hCG + FM); bST and hCG on Day 7 and S on Day 16 (Group bST + hCG), or bST and hCG on Day 7 and FM on Day 16 (Group bST + hCG + FM). The aforementioned treatments were administered at the following doses: 2.2 mg/kg FM (Banamine (R); Intervet Schering-Plough, Cotia, SP, Brazil), 500 mg bST (Boostin (R); Intervet Schering-Plough), and 2500 IU hCG (Chorulon (R); Intervet Schering-Plough). Pregnancy diagnosis was performed 40 days after TAI by transrectal ultrasonography. Pregnancy rates were not significantly different among treatments. However, there was a main effect of hCG treatment to increase pregnancy rates (63.0 vs. 55.4%; P = 0.001). Concentrations of P(4) did not differ significantly among groups on Day 7 or on Day 16. However, consistent with the higher pregnancy rates, hCG increased P(4) concentrations on Day 16 (10.6 vs. 9.6 ng/mL, respectively; P = 0.05). We concluded that hCG treatment 7 days after TAI improved pregnancy rates of lactating Nelore cows, possibly via a mechanism leading to induction of higher P(4) concentrations, or by reducing the luteolytic stimulus during maternal recognition of pregnancy. (C) 2011 Elsevier B.V. All rights reserved.

Growth hormone mRNA expression in the pituitary of Bos indicus and Bos taurus x Bos indicus crossbred young bulls treated with recombinant bovine somatotropin

The effects of breed and of recombinant bovine somatotropin (rbST) treatment on growth hormone gene expression were studied in young bulls. The experiment was completely randomized in a [2 × 2]-factorial arrangement, using two levels of rbst (0 or 250 mg/animal/14 days), and two breed groups (Nelore and Simmental x Nelore crossbred). A CDNA encoding Bos indicus growth hormone was cloned and sequenced for use as a probe in Northern and dot blot analyses. Compared to the Bos taurus structural gene, the Bos indicus CDNA was found to begin 21 bases downstream from the transcription initiation site and had only two discrepancies (C to T at position 144-His and T to C at position 354-Phe), without changes in the polypeptide sequence. However, two amino acid substitutions were found for Bubalus spp., which belong to the same tribe. The rbst treatment did not change any of the characteristics evaluated (body and pituitary gland weights, growth hormone MRNA expression level). Crossbred animals had significantly higher body weight and heavier pituitaries than Nelore cattle. Pituitary weight was proportional to body weight in both breed groups. Growth hormone MRNA expression in the pituitary was similar (P>0.075) for both breed and hormonal treatment groups, but was 31.9% higher in the pure Nelore group, suggesting that growth hormone gene transcription regulation differs among these breeds.

Effects of bovine somatotropin injection on serum concentrations of progesterone in non-lactating dairy cows

The objective of this experiment was to evaluate the effects of bovine somatotropin administration on serum concentrations of glucose, insulin, NEFA, IGF-I, and progesterone (P4) in ovariectomized non-lactating dairy cows receiving exogenous P4, as a model to estimate treatment effects on hepatic P4 degradation. Ten non-lactating, non-pregnant, and ovariectomized Gir×Holstein cows were assigned to the experiment (d -14 to 27). On d 0, cows were ranked by BW and BCS, and randomly assigned to one of two treatments: (1) bovine somatotropin (BST; n=5) or (2) saline control (control; n=5). Cows assigned to the BST treatment were administered s.c. injections containing 500. mg of sometribove zinc on d 0, 9, and 18 of the experiment, whereas control cows concurrently received a 10-mL s.c. injection of 0.9% saline. On d -2, cows were inserted with an intravaginal releasing device containing 1.9. g of P4, which remained in the cows until the end the experiment (d 27). Cow BW and BCS were assessed on d -14, 0, and 27. Blood samples were collected daily from d 0 to d 27, at 0 (immediately before), 1, and 2. h relative to concentrate feeding for determination of serum glucose, insulin, NEFA, P4, and IGF-I concentrations. Concentrations of glucose, NEFA, and insulin obtained prior to feeding (0. h) were used to determine pre-prandial revised quantitative insulin sensitivity check index (RQUICKI). No treatment effects were detected for BW (P=0.72) and BCS change (P=0.79) during the experiment. Beginning on d 2 of the experiment, BST cows had greater (P≤0.01) serum IGF-I concentrations compared with control cohorts (treatment×day interaction; P<0.01). Cows receiving BST had greater (P≤0.05) insulin concentrations compared with control cohorts from d 8 to d 11, d 16 and 17, as well as from d 19 to d 21 of the experiment (treatment×day interaction; P<0.01). Cows receiving BST had greater (P≤0.01) mean glucose and NEFA concentrations, as well as reduced (P<0.01) mean RQUICKI during the experiment compared with control cohorts. No treatment effects, however, were detected (P=0.73) for serum P4 concentrations. In conclusion, results from this experiment indicate that hepatic P4 catabolism is not directly regulated by circulating IGF-I, whereas BST administration decreases insulin sensitivity in non-lactating dairy cows in adequate nutritional status. © 2013 Elsevier B.V.